Quantitative Analysis of Pork Adulteration in Beef by Double Antibody Sandwich Enzyme-Linked Immunosorbent Assay Combined With Mathematical Models

Abstract

Adulterating pork into other meat is a global concern, and efficient methods for identifying meat species are of prime importance in quality assurance and regulation. Enzyme-linked immunosorbent assay (ELISA) is one of the most widely used methodologies for porcine speciation. Thermostable proteins were used as suitable marker proteins for species identification in heat-processed or commercial meat products. However, the continuous pursuit of higher sensitivity narrowed the analytical range of quantitation (ROQ) resulting in reduced resolution. In our study, murine monoclonal antibodies (sTn-4C10 and sTn-4B1-HRP) which are specific to skeletal muscle troponin were used as capture antibodies and detection antibodies in a double antibody sandwich ELISA (DAS–ELISA) to measure the percentage of pork in beef. When the optimal loading concentrations of sTn-4C10, sTn-4B1-HRP, and adulterated samples were set as 2, 0.2, and 20 μg/mL, respectively, this DAS–ELISA assay combined with the optimal mathematical model had an analytical ROQ of 5.79%–91.33% (w/w) in the absence of beef. The recovery rates ranged from 96% to 100%, and the coefficient of variation was below 4.5% within the ROQ. This work has widened the analytical ROQ and improved the detection resolution.